Summary
Collections of micro‐organisms are a crucial element of life science research infrastructure but are vulnerable to loss and damage caused by natural or man‐made disasters, the untimely death ...or retirement of personnel, or the loss of research funding. Preservation of biological collections has risen in priority due to a new appreciation for discoveries linked to preserved specimens, emerging hurdles to international collecting and decreased funding for new collecting. While many historic collections have been lost, several have been preserved, some with dramatic rescue stories. Rescued microbes have been used for discoveries in areas of health, biotechnology and basic life science. Suggestions for long‐term planning for microbial stocks are listed, as well as inducements for long‐term preservation.
A rapid and inexpensive method for estimating lipid content of yeasts is needed for screening large numbers of yeasts samples. Nile red is a fluorescent lipophilic dye used for detection and ...quantification of intracellular lipid droplets in various biological system including algae, yeasts and filamentous fungi. However, a published assay for yeast is affected by variable diffusion across the cell membrane, and variation in the time required to reach maximal fluorescence emission. In this study, parameters that may influence the emission were varied to determine optimal assay conditions. An improved assay with a high-throughput capability was developed that includes the addition of dimethyl sulfoxide (DMSO) solvent to improve cell permeability, elimination of the washing step, the reduction of Nile red concentration, kinetic readings rather than single time-point reading, and utilization of a black 96-well microplate. The improved method was validated by comparison to gravimetric determination of lipid content of a broad variety of ascomycete and basidiomycete yeast species.
► An improved Nile red assay for estimation of intracellular yeast lipid is developed. ► The use of 96-well microplate is high-throughput for screening large sample number. ► No prior cell washing with buffer is needed. ► Dimethyl sulfoxide improves Nile red staining for certain species. ► Kinetic reading allows different yeast strains to reach maximum emission.
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Molecular genetic study of 22 methanol-assimilating
Komagataella
strains maintained at the UCDFST collection (United States) was carried out. Multigene phylogenetic analysis revealed that strains ...previously assigned to “
Pichia pastoris
” based on the standard taxonomic tests actually belong to four different species:
K. pastoris
,
K. phaffii
,
K. mondaviorum
, and
K. ulmi
. Seven
Komagataella
species are practically indistinguishable by the standard morphological and physiological tests, and have similar D1/D2 sequences. Multigene phylogenetic analysis should be applied for reliable delineation of all seven
Komagataella
species. The biogeography of
Komagataella
yeasts is noteworthy.
K. mondaviorum
,
K. kurtzmanii
,
K. phaffii
,
K. populi
, and
K. ulmi
have been found only in North America, while
K. pastoris
and
K. pseudopastoris
are characteristic of Europe.
The subphylum Saccharomycotina is a lineage in the fungal phylum Ascomycota that exhibits levels of genomic diversity similar to those of plants and animals. The Saccharomycotina consist of more than ...1 200 known species currently divided into 16 families, one order, and one class. Species in this subphylum are ecologically and metabolically diverse and include important opportunistic human pathogens, as well as species important in biotechnological applications. Many traits of biotechnological interest are found in closely related species and often restricted to single phylogenetic clades. However, the biotechnological potential of most yeast species remains unexplored. Although the subphylum Saccharomycotina has much higher rates of genome sequence evolution than its sister subphylum, Pezizomycotina , it contains only one class compared to the 16 classes in Pezizomycotina . The third subphylum of Ascomycota , the Taphrinomycotina , consists of six classes and has approximately 10 times fewer species than the Saccharomycotina . These data indicate that the current classification of all these yeasts into a single class and a single order is an underappreciation of their diversity. Our previous genome-scale phylogenetic analyses showed that the Saccharomycotina contains 12 major and robustly supported phylogenetic clades; seven of these are current families ( Lipomycetaceae , Trigonopsidaceae , Alloascoideaceae , Pichiaceae , Phaffomycetaceae , Saccharomycodaceae , and Saccharomycetaceae ), one comprises two current families ( Dipodascaceae and Trichomonascaceae ), one represents the genus Sporopachydermia , and three represent lineages that differ in their translation of the CUG codon (CUG-Ala, CUG-Ser1, and CUG-Ser2). Using these analyses in combination with relative evolutionary divergence and genome content analyses, we propose an updated classification for the Saccharomycotina , including seven classes and 12 orders that can be diagnosed by genome content. This updated classification is consistent with the high levels of genomic diversity within this subphylum and is necessary to make the higher rank classification of the Saccharomycotina more comparable to that of other fungi, as well as to communicate efficiently on lineages that are not yet formally named.
Ten different versions of the D1/D2 divergent domain of the large-subunit ribosomal DNA were identified among interbreeding members of the yeast species
Clavispora lusitaniae. One major polymorphism, ...located in a 90-bp structural motif of the D2 domain, exists in two versions that differ by 32 base substitutions. Three other polymorphisms consist of a two-base substitution, a two-base deletion, and a single-base deletion, respectively. The polymorphisms are independent of one another and of the two mating types, indicating that the strains studied belong to a single, sexually active Mendelian population. Several strains were heterogeneous for one or more of the polymorphisms, and one strain was found to be automictic and capable of producing asci on its own by isogamous conjugation or by bud–parent autogamy. These observations suggest circumspection in the use of sequence divergence as the principal criterion for delimiting yeast species.
We previously reported the isolation of a 21.5-kb genomic DNA fragment from Pseudomonas sp. strain ADP, which contains the atzA gene, encoding the first metabolic step for the degradation of the ...herbicide atrazine (M. de Souza, L.P. Wackett, K.L. Boundy-Mills, R.T. Mandelbaum, and M.J. Sadowsky, Appl. Environ. Microbiol. 61:3373-3378, 1995). In this study, we show that this fragment also contained the second gene of the atrazine metabolic pathway, atzB. AtzB catalyzed the transformation of hydroxyatrazine to N-isopropylammelide. The product was identified by use of high-performance liquid chromatography, mass spectrometry, and nuclear magnetic resonance spectroscopy. Tn5 mutagenesis of pMD1 was used to determine that atzB was located 8 kb downstream of atzA. Hydroxyatrazine degradation activity was localized to a 4.0-kb ClaI fragment, which was subcloned into the vector pACYC184 to produce plasmid pATZB-2. The DNA sequence of this region was determined and found to contain two large overlapping divergent open reading frames, ORF1 and ORF2. ORF1 was identified as the coding region of atzB by demonstrating that (i) only ORF1 was transcribed in Pseudomonas sp. strain ADP, (ii) a Tn5 insertion in ORF2 did not disrupt function, and (iii) codon usage was consistent with ORF1 being translated. AtzB had 25% amino acid identity with TrzA, a protein that catalyzes a hydrolytic deamination of the s-triazine substrate melamine. The atzA and atzB genes catalyze the first two steps of the metabolic pathway in a bacterium that rapidly metabolizes atrazine to carbon dioxide, ammonia, and chloride
Refractory periods at the onset of dispersal occur when individuals initially do not settle in response to cues from naturally occurring habitats. The current paper focuses on relationships between ...refractory periods and habitat selection behaviour when natal dispersers use sequential search tactics. We first show how individual differences in acceptance thresholds can produce predictable relationships across individuals between the duration of their refractory periods and the time when they become responsive to cues from less preferred habitats. We then test these predictions using recurrent F1 crosses (genotypes) of
Drosophila melanogaster. We found consistent differences between genotypes with respect to the duration of their refractory periods, and strong correlations across the genotypes between the duration of refractory periods and the time when individuals accepted cues from less preferred habitats, the time when they accepted habitats with supernormal cues, and an estimate of the total time available for search. Taken together, these results support the hypothesis that individual differences in acceptance thresholds can lead to correlations across individuals between the duration of the refractory period and the time during search when they accept less preferred habitats. We discuss the implications of these results for studies of habitat selection by natal dispersers, and for studies of individual differences in other behavioural contexts (i.e. mate choice) in which animals search for a single item with a large impact on fitness.
The mission of the United States Culture Collection Network (USCCN; http://usccn.org) is "to facilitate the safe and responsible utilization of microbial resources for research, education, industry, ...medicine, and agriculture for the betterment of human kind." Microbial culture collections are a key component of life science research, biotechnology, and emerging global biobased economies. Representatives and users of several microbial culture collections from the United States and Europe gathered at the University of California, Davis, to discuss how collections of microorganisms can better serve users and stakeholders and to showcase existing resources available in public culture collections.
We previously identified a Pseudomonas sp. strain, ADP, which rapidly metabolized atrazine in liquid culture, agar plates, and soils (R. T. Mandelbaum, D. L. Allan, and L. P. Wackett, Appl. Environ. ...Microbiol. 61:1451-1457, 1995). In this study, we report the cloning and partial characterization of a gene region from Pseudomonas sp. strain ADP that encodes atrazine degradation activity. A 22-kb EcoRI genomic DNA fragment, designated pMD1, was shown to encode atrazine dechlorination activity in Escherichia coli DH5 alpha. Atrazine degradation was demonstrated by a zone-clearing assay on agar medium containing crystalline atrazine and by chromatographic methods. A gene conferring the atrazine-clearing phenotype was subsequently subcloned as a 1.9-kb AvaI fragment in pACYC184, designated pMD4, and was expressed in E. coli. This result and random Tn5 mutagenesis established that the 1.9-kb AvaI fragment was essential for atrazine dechlorination. High-pressure liquid and thin-layer chromatographic analyses were used to rigorously establish that E. coli containing pMD4 degraded atrazine and accumulated hydroxyatrazine. Hydroxyatrazine was detected only transiently in E. coli containing pMD1. This is consistent with the idea that hydroxyatrazine is the first metabolite in atrazine degradation by Pseudomonas sp. strain ADP. A 0.6-kb ApaI-PstI fragment from pMD4, containing the putative atrazine chlorohydrolase gene, hybridized to DNA from atrazine-degrading bacteria isolated in Switzerland and Louisiana. These data suggest that genes encoding atrazine hydrolysis to hydroxyatrazine are widespread in nature and contribute to the formation of hydroxyatrazine in soil, a reaction previously attributed solely to abiotic processes
Genomic information is essential for taxonomic, phylogenetic, and functional studies to comprehensively decipher the characteristics of microorganisms, to explore microbiomes through metagenomics, ...and to answer fundamental questions of nature and human life. However, large gaps remain in the available genomic sequencing information published for bacterial and archaeal species, and the gaps are even larger for fungal type strains. The Global Catalogue of Microorganisms (GCM) leads an internationally coordinated effort to sequence type strains and close gaps in the genomic maps of microorganisms. Hence, the GCM aims to promote research by deep-mining genomic data.
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